A successful biodefense strategy involves the large-scale immunization of individuals to provide protective immunity against deadly pathogens from anthrax to influenza. Such vaccines depend entirely on the generation of immunological memory mediated by long-lived memory T cells that coordinate efficacious clearance of the pathogen. Understanding the precise mechanisms for the generation, long-term maintenance and superior functional properties of memory T cells is therefore of paramount importance for the design of effective vaccines as part of a preemptive biodefense plan. My laboratory has been studying the memory immune response directed against the hemagglutinin (HA) protein of influenza. HA-specific immunity protects an individual from subsequent infection with influenza virus and is targeted in many influenza vaccine preparations. We have identified novel and profound heterogeneity within influenza HA- specific memory CD4 T cells. In particular, we have found that HA-specific memory T cells are heterogeneous for expression of a homing receptor that directs T cells to lymphoid compartments. This phenotypic heterogeneity, in turn, delineates functional subsets of HA-specific memory T cells. Here, we propose experiments to dissect the mechanisms for the generation, maintenance, in vivo homing properties, and gene expression profiles of these memory subsets. We have also identified heterogeneity in the ability of HA-specific memory T cells to produce different types of effector cytokines according to the nature of the recall stimulus. These findings reveal remarkable functional plasticity of memory T cells that has direct implications for vaccine design aimed at maintaining protective immunity, and we propose to examine mechanisms underlying this functional flexibility, including determining how bacterial superantigens such as Staphylococcal Enterotoxin B (SEB) may alter HA- specific memory T cell function. Our results suggest that pathogen-specific memory immune responses are mediated by memory T cells with diverse functional and phenotypic properties, and that the very success of vaccines to generate a long-lived anamnestic response depends on generating heterogeneous memory T cells. This research proposal will therefore address the basic mechanisms underlying memory T cell heterogeneity, using novel cellular immunologic and molecular approaches.